Modulation of Transforming Growth Factor-βProduction in Normal Human Osteoblast-Like Cells by 17β-Estradiol and Parathyroid Hormone

Abstract

Although our laboratory has reported that normal human osteoblast-like (hOB) cells contain estrogen receptors, we have failed to find major effects of 17 beta-estradiol (E2) on modulation of proliferation of bone matrix protein production by hOB cells. Because the major effect of E2 in vivo is to decrease bone resorption and because transforming growth factor-beta (TGF-beta) has been reported to decrease osteoclast-mediated bone resorption, we have tested the hypothesis that the effect of E2 on osteoclast activity is, at least in part, indirectly mediated by enhancing production of TGF-beta by osteoblasts. We therefore have extended our studies to examine possible TGF-beta gene expression including the modulation of the release of TGF-beta by E2 in near homogenous populations of hOB cells. TGF-beta protein production was measured using growth inhibition of CCL-64 cells and verified by blocking effects with anti-TGF-beta antibodies. TGF-beta 1 messenger RNA (mRNA) steady state levels were assessed by northern blot analysis and quantitated by densitometric measurement using 18S ribosomal RNA as a reference. There was an E2 dose-dependent increase in TGF-beta protein production within 24 h of challenge with E2. Northern blots from these cells demonstrated a dose-dependent increase in steady state mRNA levels of TGF-beta 1 within 6 h of treatment. PTH was also a potent stimulator of TGF-beta protein and message levels in a dose-dependent manner. Interestingly, coincubation of equimolar concentrations of E2 and PTH (10(-8) M) abrogated the stimulation of TGF-beta 1 mRNA and protein. Decreasing the relative concentration of PTH in this coincubation with E2 increased TGF-beta 1 mRNA and protein levels. These data support the fact that E2 modulates TGF-beta production in osteoblasts. In this manner TGF-beta may mediate E2 inhibition of osteoclast activity.

Parathyroid hormone increases the concentration of insulin-like growth factor-I and transforming growth factor beta 1 in rat bone.

“…Since exact figures on the cross-reactivity of the rat growth factors in our assays are unknown, it is possible that the correct absolute concentrations of these peptides may be different from the ones reported here. However (11)(12)(13)(14)(15) In the study presented here, we did not observe any increase in serum IGF-I during the course of the experiment. Based on the 95% confidence intervals of the IGF-I measurements, increases of > 10% should have been readily detected.…”

“…The role of IGF-I as mediator of the anabolic effects of PTH on bone formation is additionally supported by findings that the stimulation of collagen synthesis in cultured rat calvariae after transient PTH administration can be suppressed by antibodies to IGF-I (13). PTH has also been reported to stimulate the secretion of IGF-II in mouse calvariae cultures ( 12, 14), whereas Oursler et al observed a dose-dependent stimulation of TGF-p-bioactivity and TGF-P3-message with PTH in cultures of human osteoblast-like cells (15).…”

Estrogen‐TGFβ cross‐talk in bone and other cell types: Role of TIEG, Runx2, and other transcription factors

“…TGFβ1 is present in the greatest abundance in bone [Seyedin et al, 1985]. This family of growth factors is known to be induced by estrogen in multiple cell types including human OBs [Oursler et al, 1991a] and OCs [Oursler et al, 1994]. In spite of their high sequence homology, the in vivo functions of the three TGFβ isoforms are highly divergent as analyzed by gene knockouts.…”

“…This increase in TGFβ production then acts on the surrounding OB cells to activate the TGFβ signaling pathway. In bone, E 2 induces TGFβ synthesis in several cell types including human OB cells [Oursler et al, 1991a;Hering et al, 1995;Slater et al, 1995;Hughes, 1998;Bord et al, 2001a], human bone marrow cells [Hong et al, 2004], as well as mouse and rat OB cells [Finkelman et al, 1992;Hughes et al, 1996;Narayana Murthy et al, 2006], mouse osteocytes [Heino et al, 2002], and even chicken OCs [Robinson et al, 1996]. In many of the above studies, the levels of TGFβ gene expression or protein were measured, while in other studies TGFβ activity was determined via activation of the R-Smads.…”

“…Following the discovery of ER expression in human OBs, our laboratory identified TGFβ1 as one of the few growth factors known to be regulated by E 2 in these cells [Oursler et al, 1991a]. In order to further understand the molecular mechanisms of TGFβ action in OBs, gene expression studies were performed which identified TIEG as an early response gene to TGFβ treatment [Subramaniam et al, 1995].…”